Phase Contrast STEM for Cryo-EM
This project aims to enhance cryo-electron tomography in biology using high-resolution scanning transmission EM, improving imaging quality and enabling new insights into cellular structures.
Projectdetails
Introduction
Cryo-electron microscopy has revolutionized the field of structural biology, primarily for macromolecular structure but also for cells and tissue sections—achieving resolutions at the limit of physical optics. While wide-field transmission EM (TEM) with phase contrast by defocus is the most commonly used modality in biology, the alternative, scanning transmission EM (STEM), has emerged as the mode of choice for atomic resolution in materials science.
Objectives
Seeking to endow biology with the benefits of STEM, our lab established STEM for cryo-tomography of biological cells and demonstrated its advantages for thick specimens and compositional contrast. We now seek to extend cryo-STEM to high-resolution, with an emphasis on tomography, by means of coherent detection (Obj1). This will be achieved by the method of integrated differential phase contrast (iDPC) using a segmented detector, from which we obtain simultaneously phase and depth contrast in a single scan.
Expected Benefits
The major expected benefits are:
- Minimization of image aberration, especially defocus with its associated complications for image interpretation.
- Reduction of beam-induced radiation damage by means of flexible scan and sampling patterns.
- Improved reconstruction for tomography based on tailored data acquisition.
Validation and Applications
We will validate the new methods for single particle analysis on standard macromolecular substrates and compare them to current state-of-the-art methods. Further, we will apply the new developments in 3D imaging to explore novel large-scale structures in chromatin we observed recently by whole-cell cryo-STEM tomography using current, low-resolution methods (Obj2). Labelling with halogenated nucleotides will reveal sites of active transcription or DNA synthesis.
Conclusion
The proposed approaches’ expected broad applicability and STEM’s unrealized potential for hardware simplicity should together ensure the wide adoption of cryo-STEM methods in biology, accelerated by our dissemination efforts (Obj3).
Financiële details & Tijdlijn
Financiële details
Subsidiebedrag | € 2.499.987 |
Totale projectbegroting | € 2.499.987 |
Tijdlijn
Startdatum | 1-6-2022 |
Einddatum | 31-5-2027 |
Subsidiejaar | 2022 |
Partners & Locaties
Projectpartners
- WEIZMANN INSTITUTE OF SCIENCEpenvoerder
Land(en)
Vergelijkbare projecten binnen European Research Council
Project | Regeling | Bedrag | Jaar | Actie |
---|---|---|---|---|
4D scanning transmission electron microscopy for structural biologyThis project aims to develop advanced 4D-BioSTEM methodologies for cryo-EM to enhance contrast and resolution, enabling structure determination of small proteins and complex biological samples. | ERC Synergy ... | € 7.489.397 | 2024 | Details |
Streamlining structural biology: Developing a high-throughput cloud-based cryo-electron tomography platformDevelop a cloud-based workflow for automated high-throughput cryo-electron tomography data analysis to democratize access and reduce processing time, enhancing molecular research. | ERC Proof of... | € 150.000 | 2023 | Details |
In situ structural biology of human immune defenceDeveloping super-resolution cryo-correlative light and electron microscopy (SRcryoCLEM) to enhance in situ structural biology for antibody-mediated immunity and therapeutic advancements. | ERC Consolid... | € 1.976.553 | 2025 | Details |
Making time-resolved cryo-EM available for the community of structural biologists: validate, improve, deriskThis project aims to develop and commercialize a novel time-resolved cryo-EM sample preparation method to enhance protein structure analysis and accessibility for users. | ERC Proof of... | € 150.000 | 2024 | Details |
Advanced X-ray Energy-sensitive Microscopy for Virtual HistologyThis project aims to develop a prototype phase-contrast micro-CT scanner for non-invasive 3D histology to enhance volumetric analysis of tissue samples, particularly lung lesions. | ERC Consolid... | € 2.000.000 | 2023 | Details |
4D scanning transmission electron microscopy for structural biology
This project aims to develop advanced 4D-BioSTEM methodologies for cryo-EM to enhance contrast and resolution, enabling structure determination of small proteins and complex biological samples.
Streamlining structural biology: Developing a high-throughput cloud-based cryo-electron tomography platform
Develop a cloud-based workflow for automated high-throughput cryo-electron tomography data analysis to democratize access and reduce processing time, enhancing molecular research.
In situ structural biology of human immune defence
Developing super-resolution cryo-correlative light and electron microscopy (SRcryoCLEM) to enhance in situ structural biology for antibody-mediated immunity and therapeutic advancements.
Making time-resolved cryo-EM available for the community of structural biologists: validate, improve, derisk
This project aims to develop and commercialize a novel time-resolved cryo-EM sample preparation method to enhance protein structure analysis and accessibility for users.
Advanced X-ray Energy-sensitive Microscopy for Virtual Histology
This project aims to develop a prototype phase-contrast micro-CT scanner for non-invasive 3D histology to enhance volumetric analysis of tissue samples, particularly lung lesions.
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CryoGrid
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